Comparative Evaluation of the Cytotoxicity of Doxorubicin in BT-20 Triple-Negative Breast Carcinoma Monolayer and Spheroid Cultures.

Abstract

Three-dimensional cell culture models are increasingly adopted as preferred pre-clinical drug testing platforms, as they circumvent limitations associated with traditional monolayer cell cultures. However, many of these models are not fully characterized. This study aimed to characterize a BT-20 triple-negative breast carcinoma spheroid model and assess its susceptibility to doxorubicin in comparison to a monolayer model. Spheroids were developed using the liquid overlay method. Phenotypic attributes were analyzed by characterizing changes in size, gross morphology, protein content, metabolic activity, hypoxic status, and cell-cell junctions. The cytotoxic range of doxorubicin in monolayers was determined using the sulforhodamine B assay, and the comparative effect of toxic and sub-toxic concentrations was assessed in both spheroids and monolayers. Similar to the in vivo microenvironment, spheroids had a heterogeneous spatial cytoarchitecture, inherent hypoxia and strong adherens junctions. Doxorubicin induced dose-dependent cytotoxicity in monolayers (IC25: 130 nM, IC50: 320 nM and IC75: 1580 nM); however, these concentrations did not alter the spheroid size or acid phosphatase activity. Only concentrations ≥6 µM had any effect on spheroid integrity. In comparison to monolayers, the BT-20 spheroid model has decreased sensitivity to doxorubicin and could serve as a better model for susceptibility testing in triple-negative breast cancer.